Automated analysis of individual particles using a commercial capillary electrophoresis system

Abstract

Capillary electrophoretic analysis of individual submicrometer size particles has been previously done using custom-built instruments.<br><br>Despite that these instruments provide an excellent signal-to-noise ratio for individual particle detection, they are not capable of performing<br><br>automated analyses of particles. Here we report the use of a commercial Beckman P/ACE MDQ capillary electrophoresis (CE) instrument<br><br>with on-column laser-induced fluorescence (LIF) detection for the automated analysis of individual particles. The CE instrument was modified<br><br>with an external I/O board that allowed for faster data acquisition rates (e.g. 100 Hz) than those available with the standard instrument settings<br><br>(e.g. 4 Hz). A series of eight hydrodynamic injections expected to contain 32±6 particles, each followed by an electrophoretic separation<br><br>at −300Vcm−1 with data acquired at 100 Hz, showed 28±5 peaks corresponding to 31.9 particles as predicted by the statistical overlap<br><br>theory. In contrast, a similar series of hydrodynamic injections followed by data acquisition at 4Hz revealed only 8±3 peaks suggesting<br><br>that the modified system is needed for individual particle analysis. Comparison of electropherograms obtained at both data acquisition rates<br><br>also indicate: (i) similar migration time ranges; (ii) lower variation in the fluorescence intensity of individual peaks for 100 Hz; and (iii) a<br><br>better signal-to-noise ratio for 4Hz raw data. S/N improved for 100 Hz when data were smoothed with a binomial filter but did not reach<br><br>the S/N values previously reported for post-column LIF detection. The proof-of-principle of automated analysis of individual particles using<br><br>a commercially available CE system described here opens exciting possibilities for those interested in the study and analyses of organelles,<br><br>liposomes, and nanoparticles.